Retinol-binding protein 2 (RBP2) was originally identified more than 30 years ago as an intracellular binding protein for retinoids (vitamin A and its metabolites). RBP2 is expressed solely in the adult small intestine, primarily in the jejunum, and has been studied only from the context of its actions in dietary retinoid uptake and metabolism. We unexpectedly have identified a role for RBP2 in the maintenance of body weight, normal responses to a glucose challenge, and normal fasting hepatic triglyceride levels. When maintained solely on a control chow diet, 6?7 month-old male Rbp2-deficient (Rbp2-/-) mice accrue significantly more body weight as white adipose tissue, respond significantly less well to a glucose challenge, and possess significantly more hepatic fat than matched wild type littermate controls. These same metabolic phenotypes are observed when 2 month-old male Rbp2-/- mice are fed a high fat diet for 6 weeks or longer. Although we attempted to identify retinoid-dependent mechanisms that might account for these metabolic phenotypes, we did not detect any differences in tissue retinoic acid levels or in retinoid-regulated gene expression that could explain the metabolic phenotypes of the Rbp2-/- mice. Through biochemical studies, we established that RBP2 binds monoacylglycerols (MAGs), including both 1- and 2-arachidonoylglycerol, 2-oleoylglycerol, and 2-linoyleoylglycerol with very high affinities, comparable to that of retinol binding to RBP2. Based on these and other data, we concluded that RBP2 is a physiologically relevant MAG-binding protein. We are proposing to obtain in depth understanding of role(s) that RBP2 has in mediating enterocyte MAG metabolism. We view this as a first step towards our long-term goal of identifying the molecular processes that underlie the metabolic phenotypes we have observed in both chow fed and high fat diet fed male Rbp2-/- mice. We hypothesize that these phenotypes involve the actions of RBP2 as a MAG-binding protein. To this end, we propose 3 Specific Aims. In Specific Aim 1, we will identify non-retinoid lipids that are able to bind RBP2 and establish the structural properties of RBP2 that facilitate or prevent non-retinoid ligand binding to RBP2. In Specific Aim 2, we will elucidate RBP2-mediated biochemical pathways related to MAG uptake and metabolism within intestinal enterocytes. In Specific Aim 3, we will investigate in mice how RBP2 acts within the jejunum to affect metabolic phenotype development in response to feeding of different high fat diets consisting of either predominantly saturated or predominantly unsaturated fat and how these diets affect MAG and retinoid/carotenoid absorption and metabolism.